The objective of this project is to examine the mechanisms of generation of memory CD4 T cells, and the functional and biochemical characteristics of memory cells. The experimental system for studying CD4 memory uses adoptive transfer of primed T cells expressing a transgenic TCR (DO.11.10) into normal recipients, in which the antigen-specific cells can be followed quantitatively and purified. The specific aims of the project are the following: 1. Stimuli for the generation and maintenance of memory cells: The influence of priming conditions on the generation of memory cells will be defined, in order to establish correlations between the initial T cell expansion and differentiation and the size of the memory pool that develops subsequently. Using knockout mice lacking molecules involved in T cell activation and homeostasis (e.g., CD28, CD40L, IL-2, FasL), we will examine the role of these molecules in the development of memory cells. 2. Functional responses of memory T cells: Long-lived memory DO.11 T cells will be isolated from transfer recipients and analyzed for ex vivo responses to antigen, costimulation, and cytokines. The recall responses of these memory cells to immunogenic and tolerogenic forms of antigen will be defined. 3. Gene expression and biochemical alterations in memory T cells: We will examine the activation of selected genes and signaling pathways in memory cells to identify biochemical changes that may contribute to the survival and functional attributes of these cells. We will focus on pro- and anti-apoptotic proteins, cytokines and cytokine receptors, and intracellular biochemical intermediates that may account for the relative costimulator independence and tolerance resistance of memory cells. The physiologic importance of genes expressed in memory cells will be examined by introducing these genes into T cells by retrovirus-mediated transfer, and examining effects on memory cell generation. Thus, this proposal addresses basic issues of immunologic memory, which is fundamental to effective vaccination. Defining the stimuli and genes that control memory T cell development should lead to more rational strategies for vaccine development than the largely empirical approaches that are in common use.